A sensitive and effective strategy for the screening of α-glucosidase and its inhibitors was established based on carbon quantum dots (CQDs) and vanadium oxide nanoribbons. B,S,N co-doped CQDs (BSN-CQDs) were prepared by one-step microwave hydrothermal method using o-phenylenediamine, boric acid, and thiourea as raw materials. The prepared CQDs emitted yellow fluorescence (λem = 565 nm) under excitation of 410 nm, and the quantum yield was 15.3%. The fluorescence of CQDs was firstly quenched by the vanadium oxide nanoribbons via inner filter effect. Then, 2-O-α-D-glucopyranosyl-L-ascorbic acid (AA2G) was introduced as a substrate of α-glucosidase to produce ascorbic acid (AA). Vanadium oxide nanoribbons could be converted to V4+ by AA, which led to fluorescence recovery. After adding inhibitors (anti-diabetic drugs), the α-glucosidase activity was inhibited and the degree of fluorescence recovery was reduced. The established nanoprobe showed a response linearity to α-glucosidase in the range of 0.01–5 U mL−1, with a low detection limit of 0.003 U mL−1. This platform was applied for the screening of acarbose, with a detection limit of 0.02 μM. The constructed sensing strategy had the advantages of high selectivity, sensitivity, and was successfully used for α-glucosidase detection in serum samples and anti-diabetic drugs screening.